Cardiovascular disease (CVD) is the number one killer disease in the USA talking nearly a million lives each year. It is well established that high density lipoprotein (HDL) and its major protein constituent apolipoprotein (apo) A-l play a major role in reducing the risk of CVD. However the function of ApoA-ll, the second most abundant protein in HDL, has not been determined. Studies on apoA-ll have lead to mixed conclusions concerning the pro- or anti-atherogenicity of apoA-ll in HDL. This study will test the hypothesis that apoA-ll interacts with apoA-l in HDL in a site-specific manner to modulate its function. Furthermore, the effect is based on the relative amounts of apoA-l: apoA-ll present in a given HDL particle. In the mentored phase of this project, we will determine how the incorporation of apoA-ll affects apoA-l structure in discbidal reconstituted HDL particles. Relative changes in the solvent accessibility^ of specific apoA-l sequences in mixed particles vs apoA-| only HDL will assessed using the hydrogen deuterium exchange technique combined with mass spectrometry (HDX-MS). We will then extend our studies, in the independent phase, to locate conformational changes in apoA-l caused by apoA-ll in HDL particles from human plasma. This will be accomplished in stages. First, we will reconstitute spherical HDL particles that resemble native HDL by incorporating varying ratios of apoA-l:apoA-ll along with native HDL lipids. We will also generate native hybrid HDL particles by incorporating isolated apoA-ll into native HDL containing apoA-l only. The apoA-ll induced modifications of apoA-l solvent exposure will then be conrelated to functional properties including activation of various HDL remodeling factors including lecithin:cholesterol acyl transferase (LCAT), cholesteryl ester transfer protein (CETP), hepatic lipase, and endothelial lipase, in addition, receptor interactions with cubilin will be monitored. We anticipate that this work will provide significant new information on the role of apoA-il in lipoprotein metabolism and may suggest new therapeutic approaches for fighting CVD.